''[Instructions: IF your group has decided to redesign the PCR machine to address any major weakness discussed by your group or mentioned by others (see the Virtual Comment Board Powerpoint files on Blackboard, Lab Week 12) explain how in an additional paragraph.]''

''[Instructions: IF your group has decided to redesign the PCR machine to address any major weakness discussed by your group or mentioned by others (see the Virtual Comment Board Powerpoint files on Blackboard, Lab Week 12) explain how in an additional paragraph.]''

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Our kit will include a PCR machine similar to the OpenPCR machine. However, unlike the OpenPCR machine, ours comes assembled so it is ready to use out of the box. Furthermore, our PCR machine does not need a computer to operate. It has buttons on the LCD screen module that allow stand-alone programming and operation.

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Our kit will include a PCR machine similar to the OpenPCR machine. However, unlike the OpenPCR machine, ours comes assembled so it is ready to use out of the box. Furthermore, our PCR machine is constructed of durable, heat resistant plastic and does not need a computer to operate. It has buttons on the LCD screen module that allow stand-alone programming and operation. Our machine also has increased capacity, able to hold 36 PCR tubes at once.

OUR COMPANY: PCR Pros

LAB 6 WRITE-UP

Computer-Aided Design

TinkerCAD

[Instructions: A short summary (up to five sentences) of the TinkerCAD tool and how you used it in lab on November 20th]

We used the 3D CAD software TinkerCAD to modify the given PCR tube CAD file. We added linking structural members between all of the PCR tubes to make them into a single unit. This makes them more convenient to fill, as they create a base and are much harder to be moved, and transport, since they move as a single unit. Furthermore, we modified the directions the lids open. This minimizes the area the tube array takes up when the lids are open.

[Instructions: Show an image of your TinkerCAD PCR tube design here]

Implications of Using TinkerCAD for Design

[Instructions: A short paragraph discussing just one possible way to use TinkerCAD for something practical...like redesigning the OpenPCR machine, fluorimeter, camera holder, printing out some of the smaller plastic items on demand, etc. There are lots of possibilities...pick just ONE.]

CAD software, like TinkerCAD, is essential for rapid product design and prototyping. An example of this would be modifying the OpenPCR machine to hold 32 tubes rather than just 16. This would required a different PCR block that would take up more space than the original block. With CAD software, we can design a new PCR block and corresponding heating element and add it to the current CAD model of the OpenPCR machine. This way we can check to see if our design is viable and will fit in the current machine, or what other modifications we will need to make to accommodate our additions. After confirming that our design is viable, we can 3D print the parts we modified and add them to a current OpenPCR machine to practice assembly and display our design to interested parties (investors, etc.) without actually going into production.

Feature 1: Cancer SNP-Specific Primers

[Instructions: This information will come from the Week 9 exercises you did in lab. Your notes should be in a pdf file that is saved on Blackboard under your group.]

Background on the cancer-associated mutation

[Instructions: Use the answers from questions 3, 4, 5, and 7 to compose, in your own words, a paragraph about rs17879961]

DNA, deoxyribonucleic acid, is a polymer composed of the nucleotides (monomers) Adenine, Guanine, Cytosine and Thymine. The arrangement of these nucleotides in DNA encodes the genetic instructions that facilitates the development and operation of all organisms on earth. Since an organism is dependent upon its DNA for proper functioning, mutations in DNA can potentially have serious consequences. An example of a mutation is a single nucleotide polymorphism, or SNP, where a single nucleotide is replaced by another (and as a result the corresponding nucleotide on the the complementary DNA strand is also replaced). For example an Adenine is replaced by Cytosine (in which case the corresponding nucleotide on the other strand would go from Thymine to Guanine). In many cases an SNP will not cause any significant change in an organism's phenotype, but one instance where this does occur is an SNP of rs17879961 in chromosome 22 in humans. rs17879961 is in a tumor suppressor gene called CHEK2 that produces a protein CHK2 (checkpoint kinase 2) which helps regulate cell growth. When an SNP occurs at rs17879961 CHEK2 does not operate correctly. This greatly increases the risk of cancer in an affected individual.

Primer design

Forward Primer: 3'-ACTCACTTAAACCATATTCT-5'

Cancer-specific Reverse Primer: 5'-GGTCCTAAAAACTCTTACAC-3'

How the primers work: [Instructions: explain what makes the primers cancer-sequence specific. In other words, explain why the primers will amplify DNA that contains the cancer-associated SNP rs17879961, and will not exponentially amplify DNA that has the non-cancer allele.]

A DNA primer is a short strand of nucleotides (usually about 20 nucleotides) that is complementary to a region on a template DNA strand. Because DNA will naturally bind to complementary strands, the primer attaches itself to the region on the DNA strand to which it is complementary. In general, a 20 nucleotide-long DNA strand will only match a single region (or no region) on a DNA strand, and will thus consistently bind only to that region. However, primers that are less than 20 nucleotides long run the risk of matching more than one sequence on the template DNA strand. This is important because we use primers to "block out" a region on the DNA strand we wish to amplify. Because a primer can only bind to a template DNA strand that contains its exact complement, we can take advantage of this in PCR when we are looking for an SNP. If we create a primer that is the complement of a region of DNA with a known SNP, the primer will ONLY BIND to a template strand that contains this SNP. Furthermore, because PCR cannot function unless primers attach to the DNA template strands, we can use this to determine if an SNP is present in a given template strand. If a PCR reaction containing a template DNA strand and a primer with the complement of a cancer-associated SNP results in a large (exponential) amplification of DNA, we can conclude that this template strand contains the cancer-associated SNP. If, however, there is little or no amplification we can conclude that our primers did not attach properly, indicated that the cancer-associated SNP is not present on our template strand.

Feature 2: Consumables Kit

Included Consumables and Packaging:

In our kit, the consumables will consist of tubes containing the PCR reagents, tubes containing the SYBR Green I dye, the micropipette tips (to be used with the included micropipette) and safety equipment (gloves and eyewear). To ensure that users will be able to easily distinguish which tubes contain what, the tubes will have black markings on the lid to indicate contents as well as be color coordinated with their contents. Furthermore, SYBR Green I dye is prone to bleaching in the presence of ultraviolet light. Therefore, the tubes containing the SYBR Green I dye will be made out of a plastic that blocks ultraviolet light. Besides these modifications, the tubes will be the same as current PCR tubes and will be packaged as a unit (all of the tubes are connected by a small bar, and can be broken down into smaller units). Current micropipette and tip systems work well, so our kits will contain commercially available micropipettes and micropipette tips. The tips will be packaged in trays that stack on top of each other with the tips of trays on top partially inserting into the tips below. This unit will be vacuum packed in plastic. This conserves space and allows the tip tray assemblies to be packaged as a single unit. In addition to the above mentioned packaging, some consumables will have a resealable bag to safely store unused materials.

Feature 3: PCR Machine Hardware

[Instructions: Summarize how you will include the PCR machine in your system. You may add a schematic image. An image is OPTIONAL and will not get bonus points, but it will make your report look really awesome and easy to score.]

[Instructions: IF your group has decided to redesign the PCR machine to address any major weakness discussed by your group or mentioned by others (see the Virtual Comment Board Powerpoint files on Blackboard, Lab Week 12) explain how in an additional paragraph.]

Our kit will include a PCR machine similar to the OpenPCR machine. However, unlike the OpenPCR machine, ours comes assembled so it is ready to use out of the box. Furthermore, our PCR machine is constructed of durable, heat resistant plastic and does not need a computer to operate. It has buttons on the LCD screen module that allow stand-alone programming and operation. Our machine also has increased capacity, able to hold 36 PCR tubes at once.

Feature 4: Fluorimeter Hardware

[Instructions: Summarize how you will include the fluorimeter in your system. You may add a schematic image. An image is OPTIONAL and will not get bonus points, but it will make your report look really REALLY awesome and easy to score.]

[Instructions: IF your group has decided to redesign the fluorimeter to address any major weakness discussed by your group or mentioned by others (see the Virtual Comment Board Powerpoint files on Blackboard, Lab Week 12) explain how in an additional paragraph.]

[Instructions: This section is OPTIONAL, and will get bonus points if answered thoroughly and correctly. Here is a chance to flex some intellectual muscle. In your own words, discuss what the results for calculations 3 and 4 imply about the reliability of CHEK2 PCR for predicting cancer. Please do NOT type the actual numerical values here. Just refer to them as being "less than one" or "very small." The instructors will ask you to submit your actual calculations via e-mail. We are doing so for the sake of academic integrity and to curb any temptation to cheat.]